LACTOPHENOL COTTON BLUE METHOD

 

LACTOPHENOL COTTON BLUE METHOD

AIM: To stain the given fungi culture for the studying the morphology.

PRINCIPLE: Fungus is eukaryotic organism and they are classified into two main groups that is yeast and molds. Its cell wall is made up of chitin. Fungal cells have both macroscopic as well as microscopic structure. Lactophenol Cotton Blue reagent is used for staining as well as for wet mounting of fungi. The lactophenol cotton blue (LPCB) wet mount preparation is the most widely used method of staining and observing fungi and is simple to prepare. The preparation has three components: Lactic acid preserves the fungal structure and clears the tissue while phenol acts as a disinfectant and cotton blue imparts blue colouration to the fungal spores and hyphae.

MATERIALS REQUIRED: LPCB Stain, Fungi culture, Glass slide, Cover slip, Inoculation needle, Forceps.

PROCEDURE:

1) Place a drop of Lactophenol Cotton Blue reagent on a clean and dry slide. The stain imparts a blue colouration on hyphae.

2) By using a nichrome inoculating wire, carefully tease the fungal culture, into a thin preparation.

3) Place a coverslip on the preparation. Wait for about 5 minutes.

4) Observe first under microscope with low power for screening in low intensity.

RESULT: Fungal spores, hyphae, and fruiting structures stain blue while the background stains pale blue. For example, Aspergillus niger stains the hyphae and fruiting structures a delicate blue with a pale blue background.